Abstract: The major reasons underlying the remarkable global rise in obesity remain unclear. While, diet composition and life-style factors such as physical activity undoubtedly have important roles in determining body composition, maternal overweight (OW) status ( BMI > 25) at conception may be a critical synergizing factor. In clinical studies, we found that maternal OW status significantly increased risk of OW (i.e. the % above the 95th percentile) at 6 months of age, in infants born with normal body weights. Our overall objective is to investigate interactions in the offspring, between an obesegenic environment (consumption of a high-fat diet, HFD) and prior exposure to maternal OW during gestation. To this end, we have developed a rat model in which metabolic and endocrine abnormalities akin to obesity in human subjects were reproduced prior to conception. Exposure to maternal OW did not influence birth weight/size of offspring. However, offspring from OW dams when fed a HFD became obese to much greater extent compared to offspring from lean dams on the same diet, without changes in caloric intake. Offspring of OW rats also developed hyperinsulinemia, adipose tissue hypertrophy and showed lipogenic gene expression. Based on our preliminary studies, we hypothesize that increased glucose flux into adipose tissue and accelerated lipogenesis causes obesity in the offspring of OW dams when fed a HFD. The proposed studies are aimed at understanding the underlying differences in energy balance and insulin signaling prior to the development of obesity in the offspring. In Specific Aim 1, indirect calorimetry will be utilized to examine if gestational OW decreases energy expenditure in the offspring. In Specific Aim 2, we will test the hypothesis that maternal OW leads to skeletal muscle insulin resistance in the offspring, consequently increasing glucose partitioning into adipose tissue, lipogenesis and fat mass accretion. This will be addressed via hyperinsulinemic-euglycemic clamp experiments using labeled tracers, and will reveal the status of insulin sensitivity in the individual tissues (skeletal muscle, adipose and liver) and whole-body glucose flux. Studies examining responsiveness of adipose tissue to insulin signaling via IR-PI3K-Akt to increase lipogenic gene expression and de novo lipid synthesis and will be performed in Specific Aim 3. Finally, Specific Aim 4 will employ physical activity prior to conception as an intervention and will test the hypothesis that increased physical activity (treadmill running) in the OW dams prior to conception will improve offspring insulin sensitivity and mitigate increased adipose tissue anabolism. Overall, these studies will elucidate important metabolic mechanisms in the offspring altered by maternal OW and high-fat diets, which may contribute to increased susceptibility to obesity.